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Studies on the Absorption of Practically Water-Insoluble Drugs Following Injection VI: Subcutaneous Absorption from Aqueous Suspensions in Rats
(3)E.Secher-Hansen, H.Langgard, and J. Schou, ActaPharmacol. Toricol., 25,162,290(1967). (4)Ibid., 26.9 (1968). (5) B. E.Ballard and E. Menczel, J . Pharm. Sci., 56,1476 (1967). (6) R. H. Levy and M. Rowland, J . Pharmacokinet. Biopharm., 2, 313,337(1974). (7) R. B. Sund and J. Shou, Acts Pharmacol. Toxicol., 21, 313 (1964). (8) M. Pfeffer and D. R. Van Harken, J . Pharm. Sci., 70, 449 (1981). (9)T. Tanaka, H. Kobayashi, K. Okumura, S. Muranishi, and H. Sezaki, Chem. Pharm. Bull., 22,175 (1974). (10)J. H.Leathem, Proc. SOC.Exp. Biol. Med., 68,92(1948). (11) W. L. Honrath, A. Wolff, and A. Meli, Steroids, 2,425 (1963). (12)J. J. Nora, W. D. Smith, and J. R. Cameron, J. Pediatr., 64,547
(1964). (13) E.A. Brown, T. G . Metcalf, and L. W. Slanety, Ann. Allergy, 19, 1016 (1961). (14) R. Deanesly and A. S. Parkes, J . Physiol., 78,155 (1933). (15) J. C. Bauernfeind and H. L. Newmark, Bull. Parenter. Drug Assoc., 24,169 (1970). (16) S. Muranishi, Yakugaku Zasshi, 100,687 (1980). (17) C. L. J. Coles, K. R. Heath, M. L. Hilton, K. A. Lees, P. W. Muggleton, and C. A. Walton, J . Pharm. Pharmacol., 17,87S (1965). ACKNOWLEDGMENTS The authors thank Mrs. J. Kagawa for her technical assistance and thank Prof. M. Nakagaki, Kyoto University, for his valuable comments on the manuscript.
Studies on the Absorption of Practically Water-Insoluble Drugs Following Injection VI: Subcutaneous Absorption from Aqueous Suspensions in Rats KOICHIRO HIRANO" and H I D E 0 YAMADA Received April 8,1981,from the Shionogi Research Laboratories, Shionogi & Co., Ltd., Fukushima-ku, Osaka, 5.53,Japan. publication May 20,1981. Abstract The absorption characteristics and kinetics of practically water-insoluble drugs following subcutaneous injection of their aqueous suspensions were investigated in intact rats by the local clearance method and compared with those following intramuscular injection reported previously. The plot of the cube root of the residual fraction of the drug in the injection site uersus time gave a good linear relationship under various experimental conditions. The absorption rate constant (i)increased with decreasing particle size. This increase was remarkable in the region of mean particle diameter <2-3 pm, while i t was gradual or slight in the region above this. This phenomenon was explained by the fact that the in uiuo spreading of particles of more than -3 pm was still m,ore limited by the network of the fibrous tissues. Between j and the initial drug concentration ( C o ) or injection volume (Vo),the practically important relationship a C# Voh ( g = -0.66 and h = -0.32) could approximately he derived from the experimental results. Comparison of j values among various compounds with different solubility (C,) in saline hut with similar colloidal properties (particle size distribution and sedimentation volume) showed that a log; uersus log Cl plot gave a nearly straight line with a slope of -0.5. All the results observed for the subcutaneous absorption were similar to those for intramuscular absorption and could reasonably be explained by the kinetic model proposed for intramuscular absorption. Keyphrases Absorption, subcutaneous-from aqueous suspensions of practically water-insoluble drugs, rats 0 Water-insoluble drugssubcutaneous absorption from aqueous suspensions, rats 0 Aqueous suspensions-subcutaneous absorption of practically water-insoluble drugs, rats
Although subcutaneous administration of aqueous suspensions is very popular for practically water-insoluble drugs in preclinical animal experiments, little has been studied on their absorption mechanisms and kinetics except for the particle-size effect on pharmacological responses (1). Previously investigated (2, 3) were the absorption behaviors of subcutaneously implanted solid drugs (sphere, disk, and cylindrical shapes) and the absorption kinetic process was clarified. The intramuscular absorption characteristics of practically water-insoluble drugs from aqueous suspension were investigated in detail, 500 I Journal of Pharmaceutical Sciences Vol. 71, No. 5, May 1982
Accepted for
and a kinetic equation was proposed for the drug absorption, which was obtained empirically and was very useful (4).
In the present work, similar investigations for the subcutaneous route were done with the local clearance method in intact rats. The purpose of the present study was to clarify the relationship between the absorption rate and physicochemical properties (particle size, initial drug concentration, injection volume, drug solubility, etc. ), to express this relationship in appropriate mathematical terms, and to compare the results with those for the intramuscular route reported previously (4). The findings obtained in this study will offer a novel and useful guide for more detailed screening and preclinical testing in laboratory animals of new drugs under development. EXPERIMENTAL Materials-Azo dyes (p-aminoazobenzene, p-hydroxyazobenzene, o-aminoazotoluene, and l-phenylazo-2-naphthylamine),sulfa drugs (Nl-acetylsulfamethoxazole and sulfamethoxazole), and a steroid (betamethasone dipropionate') were used as models for practically waterinsoluble drugs. These azo dyes and sulfa drugs were the same as those used in a previous study (4).Betamethasone dipropionate was of medicinal grade and its purity was ascertained to be satisfactory through elementary analysis, melting point measurement, and TLC. Methylcellulose2 and polysorbate SO3,used as dispersing agents for preparations, were the same as those reported previously (4).All other chemicals were of analytical or reagent grade. T o compare the spreading area of injected particles, the following standard particles of different sizes were also used: two polystyrene latexes (0.721and 1.305pm in mean diameter based on number size distribution)4, polyvinyltoluene latex (2.956p ~ m )spores ~, (4.94pm)s, diSchering Corp., Bloomfield, N.d. Metolose SM-15, Shinetsu Kagaku Kogyo Co.. Ltd.,'I'okyo. Kao Atlas Co., Ltd., Tokyo. 'l'he Dow Chemical ('I)., Indianapolis. lnd 6 Coulter Electronirs, I.td.. Dunstahlc.
0022-3549/82/0500-0500$0 1.0010 @ 1982, American Pharmaceutical Association
Table I-Particle Size (&), Distribution Constant ( D ) , and Sedimentation Volume ( Vsed) of Aqueous Suspensions of Test Compounds Prepared by the Controlled Preparation Method ComDound Sulfamethoxazole N'-Acetylsulfamethoxazole p-Aminoazobenzene p-Hydroxyazobenzene o-Aminoazotoluene
1-Phenvlazo-2-naphthylamine
D,,,uma 4.0 3.5 4.2 3.6-4.1 3.9 4.0
n
Table 11-List of Data ( CO,VO,and j f o r p-Hydroxyazobenzene Suspension ") Used for Estimation of Parameters g and h, and the Values Estimated
V,,,I, cm3h
CO,mg/ml
Vn, ml
1.7 1.7 2.0 2.8-3.1 2.4 2.2
0.5 1 .o 20.0
0.50 0.50 0.50 0.50 0.05 0.10 0.50 2.00 0.050
2.9 2.6 2.7 2.5-2.7 2.7 2.5
This value 0 Mean particle diameter obtained from Rosin-Rammler plot. (obtained from Rosin-Rammler plot) shows the degree of narrowness of particle size distribution. The larger the value, the narrower the particle size distribution.
vinylbenzene latex (8.99 ~ m )and ~ ,ragweed pollen (17.95 ~ m )These ~ . are nearly spherical and commonly used as size calibrators for particle size measurements. Preparation of Test Suspensions-Unless otherwise mentioned, the vehicle composed of 0.5% (w/v) of methylcellulose, 0.005% of polysorbate 80,and 0.9% NaCl was used as a dispersion medium for the same reason mentioned previously (4). This vehicle was presaturated with the drug to be dispersed and filtered through a membrane filter6, then used for making the various preparations. To regulate particle size, all test aqueous suspensions were prepared by fractionation techniques using natural or centrifugal sedimentation. The suspensions formulated according to the controlled preparation method in which they were intended to contain particles of -4 pm in average diameter, described in the previous report (4), were used unless otherwise mentioned. Hereafter, these suspensions are referred to as controlled suspensions. Colloidal properties such as mean particle diameters (Dss),distribution constants ( n ) , and sedimentation volumes (VE.) of the test suspensions prepared by this method appear in Table I. The suspensions other than the controlled ones were prepared by similar methods with appropriate modifications. After adjustment of the drug concentration in a test suspension, it was stored a t 25' until use. The syringeability for all the suspensions was good in the ) syringe ~ used in the absorption exsame needle (25G X 2.54 ~ m and periment. Procedure of Absorption Experiment-Male Wistar albino rats (250-290 g) were used in all absorption experiments. The subcutaneous region near the center of the dorsum of intact rat was selected as a model injection site and the same local clearance method described in the previous paper (5) was employed for the present investigations. Since almost all of the drug remaining in the injection site was undissolved and unchanged, the absorption experiment adopted was considered to be appropriate for understanding the true absorption phenomena from the systems of interest. Measurement of Physicochemical Properties of Test Compounds and Suspensions-For the solub es of model drugs in 0.9% (w/v) NaCl (saline) or in pH 7.25 phosphate buffer (0.067 M Na2HP04-KH2P04) d with NaC1, previously obtained data (4) were used. The es in 2% (w/v) bovine serum albumin8 in pH 7.25 isotonic phosphate buffer were determined according to a method similar to that described in the previous paper (4). The density ( p ) of the model compound, and the sedimentation volume ( Vsed) and particle size distribution (Ds8and n ) of each test suspension were also measured by the methods described in the previous report (4). Analytical Method-Samples from Absorption Experiments-The amounts of test compounds other than betamethasone dipropionate remaining in the injection site were determined colorimetrically as described previously (4). Betamethasone dipropionate was analyzed by the HPLC method under the following conditions: H P I X apparatusg; UV detector (254 nm); columnLn,25 cm X 2.1-mm 4; mobile phase, n-hexane-isopropyl ether-ethanol-HzO (5:20:1.5:10, v/v, upper layer); flow rate (pressure), 0.43-0.46 ml/min (84 kg/cm2). Samples from Other Experiments-The concentrations of model compounds other than betamethasone dipropionate in test suspensions were analyzed spectrophotometrically as described in the previous paper (4). For the analysis of betamethasone dipropionate, the optical density was read at 243 nm after dilution with ethanol-HZ0 (l:l, v/v). The sample solutions (filtrate or supernate) for the determination of the solubilities
50.0 5.0
5.0 5.0 5.0 50.0 25.0 2.5 1.25 g = -0.656 (0.049), h
Millipore memhrane filter HA., Millipore Corp., Bedford, Mass. 7 Terumo Co., Ltd., Tokyo. Sigma Chemical Co., St. Louis, Mo. Dupont 830 I L , lo Zorbax SIL,
Comment
0.27 0.14 0.025 0.015 0.12 0.10 0.046 0.032 0.020 0.027 0.045 0.11
0.10 1.00 2.00
VO:constant
CO:constant
WO:constant
= -0.315 (0.057)
0 Controlled suspension. b The values g and h were estimated by multiple regression analysis and are given together with the standard errors in parentheses.
of azo compounds in 2% bovine serum albumin solution were assayed by a colorimetric method similar to that reported previously (4). Those of sulfa drugs were assayed as follows: One milliliter of the sample solution was thoroughly mixed with 5 ml of pH 4.7 buffer [ l N HCI-1 N CH&OONa, 3 7 (v/v)] and from this mixture the sulfa drug was extracted with 10 ml of ethyl acetate. A portion of this extract was freed of the solvent in uacuo then assayed colorimetrically after diazotization according to the method described in the previous paper (4). RESULTS A N D DISCUSSION
State of the Depot a n d D r u g Absorption Kinetics-Although the subcutaneous region is composed of networks of loosely cross-linked fibrous tissues, injected solutions form a pool (depot) with lateral spread (6,7). This was also observed in experiments using aqueous suspensions of some dyes and uniform latexes. In addition, the suspension particles were confined to such networks and their spreading was not so extensive as that of the aqueous dispersion medium (vehicle) after injection. For example, even in the case of a I-ml injection of 0.5% (w/v) of uniform latex of 0.7 pm in diameter (this size seems to be the approximate lower limit for common preparations) into the dorsal subcutaneous region of rats, the resulting lateral spreading area of the particles did not exceed -30% of that of the aqueous vehicle. This spreading area became smaller with increasing particle size. These findings clearly indicated that the subcutaneously injected particles were loosely agglomerated at the injection site. In common cases, a drug in aqueous suspension injected subcutaneously must first become dissolved in the tissue fluids surrounding the particle agglomerate, diffuse into the intercellular space (or intracellular region) of the connective tissues, and then permeate the vascular membranes until it enters the blood or lymph stream. The participation of direct absorption of the fine particles by phagocytosis or some other mechanisms can not be completely denied (8,9). However, their contribution to the drug absorption may be much smaller than that of the above mentioned mechanism uia the dissolution process except for exceedingly fine particles of water-insoluble drugs (10). It was indicated previously (11) that the connective tissue ground substance formed a diffusion barrier to the subcutaneous absorption of carbohydrates with different molecular weights in aqueous solution. The relationship between the absorption rates of a variety of compounds subcutaneously implanted in rats and their physicnchemical properties Table 111-Check of Experimentally Determined g a n d h Values Using Betamethasone Dipropionate Aqueous Suspensions Experiment Number 1"
26 6
j , hr-'
CO,mg/ml
Vn, ml
2 2 1 10
0.05 0.50 0.50 0.50
j ( i ) / J ( 1) Calculated Experimental
0.48
0.49 (0.03)
0.2'2
0.26(0.03)
a Vehicle, 0.01% (w/v) polysorbate 80 + 0.01% (w/v) henzalkonium chloride + 0.38% (w/v) NaHzP042H20 1.79%(w/v)Na2HPOr12H,O; D, ( n ) ,7.1 pm ( 2 . 5 ) . Vehicle,0.5% Iw/vl methvlcellulose SM-15 0.01%( w k ) aolvsorhate 80 t 0.9% (w/v) NaCI; D.. ( n ) ,2.5 pm (2.6).
+
+
Journal of Pharmaceutical Sciences I 501 Vol. 71, No. 5, May 1982
A
B
s 0.6
-
0.4
-
+\ +\
I
I
1
0.
Figure 1-Time course of p-hydroxyazobenzene absorption from subcutaneous injection site. Each point represents the mean of a t least four experiments; vertical bar shows the standard deuiation. CO= 1 mglml; VO= 0.5 ml. were studied (2). It was concluded that the absorption from this system was solution rate limited. Visual observations of present experimental systems supported this conclusion, since the tissue fluid colored by the dissolution of a dye was almost limited to the narrow region close to the surface of the particles or their agglomerate except for a short period immediately after injection. On the basis of the above findings, it is assumed that the absorption is controlled by diffusion from the surface of the agglomerate. The process of this absorption seems to have a resemblance to that observed after intramuscular injection. Thus, the absorption model proposed for the intramuscular injection in a previous study (4) is expected to be applicable to the subcutaneous injection as well. From that model the following kinetic equations could be derived to describe the intramuscular absorption process (4). The fraction of the drug amount ( W )remaining a t the injection site to the dose ( W O was ) given as a function of time ( t ) : ( W/Wo)1l3= 1 - j t
(Eq. 1)
where j was defined as an absorption rate constant. This j could be related to the in uiuo solubility ( C s ) ,density of the drug crystal ( p ) , in uiuo dissolution rate constant ( k ) ,and the dose ( WO), using a correction parameter ( t ) for the effective surface area of the particle agglomerate for the in uiuo dissolution: j = k ~ C , p -W ~ /0~- ~ f ~ / 3
(Eq. 2)
In this equation the parameter t was assumed tobe an unknown function of the effective particle size ( D ) ,initial volume concentration (Colp) of the suspended drug, injection volume (Vo), and other factors ( U )containing hydrodynamic factors (injection speed and pressure) and the histological and physiological factors in the injection site: = F ( D , C o l p , Vo,
u)
(Eq. 3)
intramuscular absorption were checked to ascertain their applicability for the subcutaneous absorption and then developed to gain greater understanding of its phenomena. Ti m e Course of D r u g Absorption-First, the validity of Eq. 1 was checked experimentally. Figure 1 shows a plot of the residual fraction (WIWO)or its cube root of the drug remaining in the injection site against time after dorsal subcutaneous administration of the controlled aqueous suspension of p-hydroxyazobenzene into intact rats. The initial drug concentration (CO)and injection volume (Vo) are given in the legend. The cube root plot (B) gave a good linear relationship while the other (A) showed an upward curvature. This tendency was common throughout the following absorption experiments of different conditions, and the correlation coefficient obtained by the least-squares linear regression from the cube root plot was always larger than that from the linear plot. These results suggest that Eq. 1 is appropriate for describing the time course of the subcutaneous drug absorption from aqueous suspensions. In addition, the moderate magnitude of the standard deviations shown in Fig. 1demonstrated that the other factors ( U )in the parameter t might be well-controlled under the experimental conditions in this study. Effect of Particle Size on Absorption-The particle size of a suspension is an important factor for intramuscular (12-14) and subcutaneous absorption (1)as well as for absorptions from other routes (15). If the suspension particles are dispersed at the injection site too extensively to aggregate with each other, and, hence, the drug transport from the surface of each particle takes place independently, the particle size effect on the absorption may appear most strongly. tn such a case, theoretically, E = 6( W ~ / P ) ' / ~for / Dmonodispersed spherical particles (diameter, D ) and therefore is inversely proportional to the particle size (4). However, this is expected to he rare in practice because of the agglomerate formation in the injection site, except in an exceedingly dilute suspension with very fine particles. Thus, the relationship between the drug absorption
The relationship between t (or j ) and Co, VO,or D was investigated experimentally (4). In the present study, the above kinetic relationships proposed for the
0.14
-
0.10
-
0.06
-
HOURS
L.
S .1
0.02
1 I
0
'fy-.*I
I
I
4
8
12
1. 1~
PARTICLE SIZE, Dss,Mrn
F i gure 2-Effect of particle size on subcutaneous absorption of phydroxyazobenzene. Each point represents the mean of a t least three experiments; uertical bar indicates the standard deuiation. Ca = 5 rnglml; V O= 0.5 ml. 502 1 Journal of Pharmaceutical Sciences Vol. 71, No. 5, May 1982
F i g u r e 3-Relation between absorption rate constant (j) and particle size (Dss).The ualue of j was estimated by the least-squares method and is plotted with the standard error. Key: -0-, sc (VO= 0.5 ml);-0-, im (Vo = 0.05 ml);p-hydroxyazobenzene concentration of both injections, 5 mglml.
of Subcutaneous Absorption Rate Constants ( j ) with In Vitro Solubilities (6,a n d Ci) a n d Values of
Table IV-Comparison MW-0.5p-0.34
~
~
~~~~
I n Vitro Solubility C,, mg/mla C,, mg/mlb
Compound Sulfamethoxazole
5.7' 0.076 0.049 0.034 0.007 0.0003
N1-Acetylsulfamethoxazole p -Aminoazobenzene p-Hydroxyazobenzene 0 - Aminoazotoluene 1-Phenylazo-2-naphthylamine
j , hr-'
MW-O.5,,-0.34 c
7.2 <0.76 0.41 <0.69 <0.14 0.076
0.0549 0.0522 0.0672 0.0637 0.0625 0.0585
0.374 (0.015) 0.168 (0.013) 0.0652 (0.0037) 0.0420 (0.0035) 0.0193 (0.0016) 0.00241 (0.00014)
a Solubility in saline [0.9% (w/v) NaCl aqueous solution] a t 37". * Solubility.in 2% (w/v) bovine serum albumin in pH 7.25 isotonic phosphate buffer at 37". MW, molecular weight; p ! density. This term is defined in detail in the text. d Value estlmated by the least-squares method from the data shown in Fig. 6;listed with the standard error in arentheses. e Solubility in pH 7.25 isotonic phosphate buffer at 37" (solubility in saline at 37', 0.61 mg/ml). Solubilities of other test compounds in saline agreed well wit[ those in the buffer.
rate and particle size seems to be very complicated. A previous study on the intramuscular absorption showed that the increase in the absorption rate constant 0')with decreasing mean particle diameter (D,,) was remarkable in the region of D,,smaller than 2-3 pm, while it was gradual or slight in the region above this (4). Figure 2 compares subcutaneous absorption time profiles of p-hydroxyazobenzene from three preparations with different mean particle diameters (DSs)but with similar distribution constants ( n )and sedimentation volumes (Vaed):D , ( n ) ,7.4pm (2.5), 2.5pm (2.8),and 1.1 pm (2.9);V e d , 2.8-3.2cm3/g. The absorption rate constant 0') was obtained from the slope of each linear relationship in this figure and then plotted against D, in Fig. 3.For comparison, the results from similar experiments of the intramuscular route (m. gastrocnemius) are also shown. Figure 3 suggests that the relationship between j and D,, for the subcutaneous route is qualitatively similar to that for the intramuscular route. The particles smaller than 2-3 pm in diameter seem to pass more easily through the network of the fibrous tissues accompanying the spreading of the dispersion medium during injection. This may cause a looser agglomeration of the particles and result in the abrupt change in J below this particle size. Figure 4 shows the effect of particle size on the lateral spreading of the particle agglomerate. Here,six water-insoluble standard particles with different mean diameters (D, obtained from the number size distribution curve) were used for comparison. The similarity of the form of this curve to those in Fig. 3 may support the above speculation. The participation of direct absorption mechanisms involving phagocytosis must be also considered but their contributions seem to be relatively small under the present experimental conditions. In any event, the tendency of the particle size effect appears to have a very important practical meaning. Effect of Initial Dr u g Concentration and Injection Volume-If the injected particles form no agglomerate and the drug transport from each particle proceeds independently, and if this process is a rate-determining step, the absorption rate constant 0')does not depend on the dose and is inversely proportional to the particle size as mentioned previously. However, this was not the case for the subcutaneous injection of ordinary aqueous suspensions. The absorption rate constant G), given
by Eq. 2,is a function of the dose Wo and the parameter c. The term WO is the product of the initial drug concentration (CO)and injection volume ( V O ) The . parameter c is assumed to depend on COand Vo, but their relationship is not known. Therefore, the quantitative relationship between j and COor V0 is also not clear. To elucidate this relationship, the absorption time courses of the controlled suspensions of p-hydroxyazobenzene with different Co and V Owere followed, then their absorption rate constants 0') were obtained from the same plot as Fig. 1B.In Fig. 5 j is plotted against COa t fixed VO (0.5 ml) or against VOa t fixed CO(5mg/ml) on a log-log scale. The linear relationships of both plots allow the following tentative and approximate representation: where f is a constant that depends on the drug and its suspension preparation. Thus, the experimental values of g and h were estimated by multiple regression analysis from 12 sets of data for the controlled suspensions of p-hydroxyazobenzene. The original data used for this estimation and resulting g and h values are summarized in Table 11. Next, to ascertain the applicability of these experimental values (g = -0.66 and h = -0.32), additional examinations were done using aqueous suspensions of betamethasone dipropi'onate. Employing Eq. 4 and the g and h values determined above. the ratio of j ( i ) at Co(i) and Vo(i)to j(1) at Co(1) and Vo(1) for suspensions with the same drug and similar colloidal properties can be represented as:
j ( i ) / j ( I )= [Co(i)/Co(1)l-066 [ V o ( i ) l V 0 ( 1 ) ]32- ~
V,, ml 0.1
-
L '
0.1
-
0.05
-
L
.c
;.
0.5
0.2 I
I
1
1
I
1 1 1 . 1 1
I 0
1
5
10
15
MEAN PARTICLE (D), pm
20
Figure 4-Effect of particle size on the lateral spreading area o f the particle agglomerate formed by subcutaneous injection of aqueous suspension. Each point represents the mean of three experiments; vertical bar indicates the standard deuiation. These data were obtained using water-insoluble standard particles of uarious sizes under the following conditions: CO= 5 mglml; VO = 1.0 ml; measurement, 5 min after injection.
0.01
L r
2 I
5 I
1
1
1
1
I
,
-\
0.021 0.6
(Eq. 5)
Table I11 compares the calculated and experimental ratios j(i)/j(l)for the case where Co(i)/Co(l)= 10 or Vo(i)/Vo(l)= 10.These comparisons showed good mutual agreements, despite the fact that the vehicle and the colloidal properties of the suspensions tested here differed considerably from those of the controlled suspension of p-hydroxyazobenzene used for the estimation ofg and h values. Therefore, Eq. 5,obtained ex0.05
Q
(Eq. 4)
j = fC&Voh
' ' 0.5
I
t " 1 '
1
2
5
10
20
I
1
,
50
C,. mglrnl
Figure 5-Relation between absorption rate constant 6) and injection volume (Vo) or initial drug concentration (Co) for t h e controlled suspension of p-hydroxyazobenzene. T h e value of j U I Q S estimated by the least-squares method. Key: - 0 - , constant VO(0.5 ml);-0-, constant CO(5mglml). Journal of Pharmaceutical Sciences I 503 Vol. 71, No. 5. May 1982
HOURS
- 10
1.
lo-'
.
-1
"
"
L
L
r
c
..
'\
- lo-'
.
-
- /oO0
10-2
-
lo-'
lo-'
IO-~
lo-*
10
1
Ci,mg/ml
Figure 6-Comparison of absorption rate among uarious compounds (controlled suspension). Each point represents the mean of three or four experiments; vertical bar indicates the standard deuiation. Co = 5 mglml; VO = 0.5 ml. Key: -@-, I-phenylazo-2-naphthylamine; -A-, o-aminoazotoluene; p-hydroxyazobenzene; -0-, paminoazobenzene; -A-, N'-acetylsulfamethoxazole; -0-, sulfamethoxazole.
Fi g u r e 7-Relationship between absorption rate constant (j) a nd in vitrosolubility (Cs').The ualue of j was estimated by the least-squares method from the data shown in Fig. 6. Key: -@-, sc (C, = 5 mglml; vo = 0.5 ml); -0-, im (Co = 5 mglml; Vo = 0.05 ml), cited from a previous study ( 4 ) .
perimentally, is expected to be generally applicable for aqueous suspensions other than the model ones tested here. Using the above results, the relationship between the parameter t and COor VOcan be clarified empirically in the following manner:
As discussed previously, the absorption is inferred to be a diffusioncontrolled process. Diffusion coefficients of compounds having low molecular weight (MW) are known to be inversely proportional to the square root of MW (16). The in uiuo dissolution rate constant ( k ) in Eq. 9 is considered to be proportional to the diffusion coefficient, like that in uitro (2,17). Therefore, Eq. 9 can be converted to
-+-,
j = fCO-O66VO-0.32
(Eq. 6)
Employing the relationship WO= CoVo, Eq. 2 can be rewritten as:
j =
(Bd/3)MW-0.5p-0.34C,Co-0.66V
-0.32
(Eq. 11)
where B is a constant. Thus, the values of MW-o.5p-o.34for model drugs were calculated and are in Table IV as the correction term. However, From the equivalency of Eqs. 6 and 7, the parameter t must satisfy the these values were similar to each other for the compounds tested here. following relationship: In contrast, the in uitro solubility was a more dominant factor and had a positive relation with the absorption rate constant. c = F(D, Colp, VO,U ) = ~ ( C O / ~ ) - ~ . ~(Eq. ~ V8 )~ ~ . ~Equation ~ 11 means that the plot of j (or J,,,, = jMW0.5e0.34,if the correction for MW and p is necessary) against in uiuo solubility (C,)'on where 6 is a term that depends on the remaining factors D and U . a log-log scale gives a straight line with a slope of unity a t the constant Equation 8 means that the parameter t depends little on the injection initial drug concentration (CO)and injection volume (VO).The closed volume ( V O )but decreases by increasing initial drug concentration (CO). circles in Fig. 7 show this plot using in uitro solubility (C',) in Table IV This also implies that the degree of agglomeration may tend to increase instead of in uiuo solubility (Cs). This gave a nearly straight line with a with an increase in Co. Substitution of Eq. 8 for c in Eq. 7 yields: slope of 0.53 (regression equation, log j = 0.525 log Cb - 0.578; correlation coefficient, r = 0.956). Using j,,,, in place of j , a similar relationship was j = (kC,j/3)p-0.34CO-0.66VO-0.32 (Eq. 9) also obtained (log jcorr = 0.534 log C', 0.660, r = 0.953). The above empirical equation obtained for the subcutaneous route In Fig. 7, similar data for the intramuscular administration (m. gasdemonstrates a great similarity to that for the intramuscular route, which trocnemius), cited from a previous study (4), are also plotted with open was given by the following equation (4): circles. Remarkably, the slopes of these plots for the two routes were almost identical. This shows a strong similarity in the absorption mechaj , = (k,C,6,/3)p-0.45Co-0.55V~-o 32 (Eq. 10) nism between the two routes. The deviation of the slope from unity obIt should be noted that the correlation between the absorption rate served in Fig. 7 may he attributed mainly to a gradual increase in the ratio constant and injection volume is identical for the two routes. C,/C, with decreasing C; caused by the relatively larger. solubilization Although Eqs. 9 and 10 are derived empirically, they should be very effect of the protein components in the body fluid for the drug having important and useful since they give relationships between the dose and a smaller solubility. The comparison of C', and C ; in Table IV supports absorption rate constant which have been unknown until now. this explanation. Comparison of Absorption R a t e among Various CompoundsThe good linear relationship between log j (or log j,,,,) and log C; Equation 9 shows that the absorption rate constants (j) of different mentioned previously is expected to be applicable for predicting the compounds depend upon the values of k , C,, and p when the preparasubcutaneous absorption rates of other compounds in the controlled tion-dependent parameters (6) are the same. As defined previously, p , suspensions from C,. For instance, the prediction (from the regression k , and C, represent the density of the drug, the in uiuo dissolution rate equation, log j,,, = 0.534 log C; 0.660) for the test material, betamconstant, and in uiuo solubility in the injection site, respectively. Of these ethasone dipropionate (MW = 504; p = 1.23 g/cm3; C, = 0.002 mg/ml) factors, only the value of p can be determined by an in uitro experiment; gave a value of 0.007 hr-I for j (at CO= 5 mg/ml and VO= 0.5 ml), which the other two ( k and C,) cannot be completely estimated from such an almost agreed with the experimental value of 0.0096 (f0.0011) hr-' obexperiment. From a practical point of view, an attempt was made to served using the following controlled suspension: D,,= 4.3 pm; n = 2.6. clarify the quantitative relationship between j and the above three factors This satisfactory result suggests that similar prediction will be possible for other drugs. (or their first approximations). Figure 6 compares the subcutaneous absorption time profiles of six test Comparison of Absorption Rates between Subcutaneous a n d compounds. In this comparison, the controlled suspensions with similar I n t r am u scu l ar Routes-Hitherto, drug absorption kinetics and colloidal properties (Table I) were used to minimize the variation in 6 in mechanisms from aqueous suspensions in the subcutaneous route have Eq. 9. All the compounds gave good linear relationships with different been clarified, and their strong similarity to those by the intramuscular slopes. Their observed absorption rate constants (j) are summarized in route, discussed previously (4), have been pointed out. A direct comTable IV with their physicochemical factors such as in uitro solubilities parison of absorption rates between the two routes seems to be usable but there has been little available data (18). Table V shows this comin saline and 2% (w/v) bovine serum albumin (C, and C;, respectively). j = kC,1'-2/3~C~-1/3V~-1/3/3
(Eq. 7)
+
+
504 I Journal of Pharmaceutical Sciences Vol. 71, No. 5,May 1982
Table V-Comparison of Absorption Rate Constants ( j ) between Subcutaneous (sc) and Intramuscular (im) Routes a j , hr-I
Compound Sulfamethoxazole p-Aminoazobenzene p-Hydroxyazobenzene o-Aminoazotoluene 1-Phenylazo-2-naphthyl-
sc
imc
0.78 0.14 0.090 (0.12 f 0.02)d 0.040 0.0050
1.10 f 0.07 0.18 f 0.01 0.17 f 0.01 0.093 f 0.005 0.0093 f 0.0006
smine
*
Controlled suspension.CO,5 mg/ml; VO,0.05 ml. Estimated by extrapolation of data shown in Table IV usin Eq 5 Experimental data (with standard error) cited from the previous reportT4). d Experimentalvalue (with standard error). parison using five controlled suspensions. To compare a t the same drug , values estimated by concentration (CO)and injection volume ( V O )the extrapolation of the data shown in Table IV using Eq. 5 were used for the absorption rate constants 0’) in the subcutaneous route. This comparison shows that the absorption rate from the subcutaneous route is slower than that from the intramuscular route for all the test suspensions. A similar tendency was previously observed for injections of drug-oil solutions (5). The relationship between j and COin the subcutaneous route differed slightly from that in the intramuscular route (Eqs. 9 and 10). Therefore, it should be noted that the difference in j shown in Table V may increase with increasing CO. REFERENCES (1) V. G. Foglia, J. C. Penhos, and E. Montuori, Endocrinology, 57, 559 (1955). (2) B. E. Ballard and E. Nelson, J. Pharmacol. Erp. Ther.,135,120 (1962). (3) B. E. Ballard and E. Nelson, J. Pharm. Sci., 51,915 (1962).
(4) K. Hirano, T. Ichihashi, and H. Yamada, Chem. Pharm. Bull., 29,817 (1981). (5) K. Hirano, T. Ichihashi, and H. Yamada, J . Pharm. Sci., 71,495 (1982). (6) E. A. Brown, T. G. Metcalf, and L. W. Slanetz, Ann. Allergy, 19, 1016 (1961). (7) F. L. Ashley, S. Braley, T. D. Rees, D. Goulian, and D. L. Ballantyne, Jr., Plastic Reconstruc. Surg., 39.411 (1967). (8) C. R. Beresford, L. Golberg, and J. P. Smith, Br. J.Pharmacol., 12, 107 (1957). (9) T. D. Rees, D. L. Ballantyne, Jr., I. Seidman, and G. A. Hawthorne, Plastic Reconstruc. Surg., 39,402 (1967). (10) J. Lewin and F. Huidobro, Acta Physiol. Lat. Am., 3, 17 (1953). (11) E. Secher-Hansen, H. L. Langgard, and J. Schou, Acta Pharmacol. Toxicol., 26,9 (1968). (12) F. H. Buckwalter and H. L. Dickison, J . Am. Pharm. Assoc. Sci. Ed., 47,661 (1958). (13) L. G. Miller and J. H. Fincher, J . Pharm. Sci., 60,1733 (1971). (14) N. Kitamori, S. Kawaziri, and T. Matsuzawa, “Abstracts of Papers,” 93th Annual Meeting of Japan Pharmaceutical Society, Tokyo, April 1973, p. 266. (15) J. H. Fincher, J . Pharm. Sci., 57,1825 (1968). (16) W. D. Stein, “The Movement of Molecules across Cell Membranes,” Academic, New York, N.Y. 1967, p. 67. (17) W. Nernst, 2. Phys. Chem., 47,52 (1904). (18) J. J. Nora, W. D. Smith, and J. R. Cameron, J . Pediatr., 64,547 (1964). ACKNOWLEDGMENTS The authors thank Mrs. J. Kagawa for her technical assistance and Prof. M. Nakagaki, Kyoto University, for his valuable comments on the manuscript.
Dissolution and Bioavailability Studies of Whole and Halved Spstained-Release Theophylline Tablets KEITH J. SIMONSX*,EVELYN M. FRITH*, and F. ESTELLE R. SIMONS$ Received July 24,1980, from the ‘Faculty of Pharmacy, and f Section of Allergy and Clinical Immunology, Department of Pediatrics, Facultv Accepted for publication August 5, 1981. of Medicine, University of Manitoba, Winnipeg, Manitoba, Canada, R3T 2N2. Abstract 0 In dissolution studies of whole and halved 100-mg sustained-release theophylline tablets, drug release from halved tablets was significantly higher. These differences were not reflected in the bioavailability studies. The area under the curve (AUC) mean absorption time and fraction-of-dose recovered in urine at 24 hr were not significantly different following the ingestion of whole or halved 100-mg tablets. The elimination rate constant, half-life, volume of distribution, plasma, and renal clearance values were consistent with values reported previously. Discrepancies were found in the 24-hr metabolite distribution as compared to literature values and may be accounted for by the age and health of the subjects and the frequency of dosing.
phylline tablets’ were used to study the effect of halving tablets on dissolution and bioavailability. No published information about the dissolution of these tablets was available. After oral administration of the 100-mg tablet, however, 90% of the dose was absorbed within 14 hr and almost 100% was absorbed by 28 hr (1). When 300-mg tablets were dissolved, 50% of the dose entered solution by 2 hr and > 90% of the dose entered solution by 6 hr (1).
Keyphrases 0 Dissolution-whole and halved sustained-release theophylline tablets Sustained-release system-dissolution of whole and halved theophylline tablets 0 Bioavailability-whole and halved sustained-release theophylline tablets 0 Theophylline-bioavailability and dissolution study of whole and halved sustained-release tablets
EXPERIMENTAL
Breaking sustained-release theophylline tablets in half is commonly practiced to achieve more accurate milligrams per kilogram dosing in children. The extent to which this affects dissolution and bioavailability is unknown. In this investigation, 100-mg sustained-release theo0022-354918210500-0505$01.0010 @ 1982. American Pharmaceutical Association
Dissolution-The official USP dissolution apparatus was used (2). Simulated gastric and intestinal fluids were used as dissolution media (2). Simulated gastric fluid, USP (Z),was prepared by dissolving 2 g of sodium chloride and 3.2 gof pepsin in 7 ml of HCI and diluting the solution to 1000 ml with distilled water. This test solution had a pH of 1.2, Simulated intestinal fluid, USP (2), was prepared hy diswlv~ng6.8 g Theo-Dur, Astra 1M4.
Pharmaceuticals Canada Ltd.. Mississauya, Canada IAX
Journal of Pharmaceutical Sciences I 505 Vol. 71,No. 5,May 1982
(1964). (13) E.A. Brown, T. G . Metcalf, and L. W. Slanety, Ann. Allergy, 19, 1016 (1961). (14) R. Deanesly and A. S. Parkes, J . Physiol., 78,155 (1933). (15) J. C. Bauernfeind and H. L. Newmark, Bull. Parenter. Drug Assoc., 24,169 (1970). (16) S. Muranishi, Yakugaku Zasshi, 100,687 (1980). (17) C. L. J. Coles, K. R. Heath, M. L. Hilton, K. A. Lees, P. W. Muggleton, and C. A. Walton, J . Pharm. Pharmacol., 17,87S (1965). ACKNOWLEDGMENTS The authors thank Mrs. J. Kagawa for her technical assistance and thank Prof. M. Nakagaki, Kyoto University, for his valuable comments on the manuscript.
Studies on the Absorption of Practically Water-Insoluble Drugs Following Injection VI: Subcutaneous Absorption from Aqueous Suspensions in Rats KOICHIRO HIRANO" and H I D E 0 YAMADA Received April 8,1981,from the Shionogi Research Laboratories, Shionogi & Co., Ltd., Fukushima-ku, Osaka, 5.53,Japan. publication May 20,1981. Abstract The absorption characteristics and kinetics of practically water-insoluble drugs following subcutaneous injection of their aqueous suspensions were investigated in intact rats by the local clearance method and compared with those following intramuscular injection reported previously. The plot of the cube root of the residual fraction of the drug in the injection site uersus time gave a good linear relationship under various experimental conditions. The absorption rate constant (i)increased with decreasing particle size. This increase was remarkable in the region of mean particle diameter <2-3 pm, while i t was gradual or slight in the region above this. This phenomenon was explained by the fact that the in uiuo spreading of particles of more than -3 pm was still m,ore limited by the network of the fibrous tissues. Between j and the initial drug concentration ( C o ) or injection volume (Vo),the practically important relationship a C# Voh ( g = -0.66 and h = -0.32) could approximately he derived from the experimental results. Comparison of j values among various compounds with different solubility (C,) in saline hut with similar colloidal properties (particle size distribution and sedimentation volume) showed that a log; uersus log Cl plot gave a nearly straight line with a slope of -0.5. All the results observed for the subcutaneous absorption were similar to those for intramuscular absorption and could reasonably be explained by the kinetic model proposed for intramuscular absorption. Keyphrases Absorption, subcutaneous-from aqueous suspensions of practically water-insoluble drugs, rats 0 Water-insoluble drugssubcutaneous absorption from aqueous suspensions, rats 0 Aqueous suspensions-subcutaneous absorption of practically water-insoluble drugs, rats
Although subcutaneous administration of aqueous suspensions is very popular for practically water-insoluble drugs in preclinical animal experiments, little has been studied on their absorption mechanisms and kinetics except for the particle-size effect on pharmacological responses (1). Previously investigated (2, 3) were the absorption behaviors of subcutaneously implanted solid drugs (sphere, disk, and cylindrical shapes) and the absorption kinetic process was clarified. The intramuscular absorption characteristics of practically water-insoluble drugs from aqueous suspension were investigated in detail, 500 I Journal of Pharmaceutical Sciences Vol. 71, No. 5, May 1982
Accepted for
and a kinetic equation was proposed for the drug absorption, which was obtained empirically and was very useful (4).
In the present work, similar investigations for the subcutaneous route were done with the local clearance method in intact rats. The purpose of the present study was to clarify the relationship between the absorption rate and physicochemical properties (particle size, initial drug concentration, injection volume, drug solubility, etc. ), to express this relationship in appropriate mathematical terms, and to compare the results with those for the intramuscular route reported previously (4). The findings obtained in this study will offer a novel and useful guide for more detailed screening and preclinical testing in laboratory animals of new drugs under development. EXPERIMENTAL Materials-Azo dyes (p-aminoazobenzene, p-hydroxyazobenzene, o-aminoazotoluene, and l-phenylazo-2-naphthylamine),sulfa drugs (Nl-acetylsulfamethoxazole and sulfamethoxazole), and a steroid (betamethasone dipropionate') were used as models for practically waterinsoluble drugs. These azo dyes and sulfa drugs were the same as those used in a previous study (4).Betamethasone dipropionate was of medicinal grade and its purity was ascertained to be satisfactory through elementary analysis, melting point measurement, and TLC. Methylcellulose2 and polysorbate SO3,used as dispersing agents for preparations, were the same as those reported previously (4).All other chemicals were of analytical or reagent grade. T o compare the spreading area of injected particles, the following standard particles of different sizes were also used: two polystyrene latexes (0.721and 1.305pm in mean diameter based on number size distribution)4, polyvinyltoluene latex (2.956p ~ m )spores ~, (4.94pm)s, diSchering Corp., Bloomfield, N.d. Metolose SM-15, Shinetsu Kagaku Kogyo Co.. Ltd.,'I'okyo. Kao Atlas Co., Ltd., Tokyo. 'l'he Dow Chemical ('I)., Indianapolis. lnd 6 Coulter Electronirs, I.td.. Dunstahlc.
0022-3549/82/0500-0500$0 1.0010 @ 1982, American Pharmaceutical Association
Table I-Particle Size (&), Distribution Constant ( D ) , and Sedimentation Volume ( Vsed) of Aqueous Suspensions of Test Compounds Prepared by the Controlled Preparation Method ComDound Sulfamethoxazole N'-Acetylsulfamethoxazole p-Aminoazobenzene p-Hydroxyazobenzene o-Aminoazotoluene
1-Phenvlazo-2-naphthylamine
D,,,uma 4.0 3.5 4.2 3.6-4.1 3.9 4.0
n
Table 11-List of Data ( CO,VO,and j f o r p-Hydroxyazobenzene Suspension ") Used for Estimation of Parameters g and h, and the Values Estimated
V,,,I, cm3h
CO,mg/ml
Vn, ml
1.7 1.7 2.0 2.8-3.1 2.4 2.2
0.5 1 .o 20.0
0.50 0.50 0.50 0.50 0.05 0.10 0.50 2.00 0.050
2.9 2.6 2.7 2.5-2.7 2.7 2.5
This value 0 Mean particle diameter obtained from Rosin-Rammler plot. (obtained from Rosin-Rammler plot) shows the degree of narrowness of particle size distribution. The larger the value, the narrower the particle size distribution.
vinylbenzene latex (8.99 ~ m )and ~ ,ragweed pollen (17.95 ~ m )These ~ . are nearly spherical and commonly used as size calibrators for particle size measurements. Preparation of Test Suspensions-Unless otherwise mentioned, the vehicle composed of 0.5% (w/v) of methylcellulose, 0.005% of polysorbate 80,and 0.9% NaCl was used as a dispersion medium for the same reason mentioned previously (4). This vehicle was presaturated with the drug to be dispersed and filtered through a membrane filter6, then used for making the various preparations. To regulate particle size, all test aqueous suspensions were prepared by fractionation techniques using natural or centrifugal sedimentation. The suspensions formulated according to the controlled preparation method in which they were intended to contain particles of -4 pm in average diameter, described in the previous report (4), were used unless otherwise mentioned. Hereafter, these suspensions are referred to as controlled suspensions. Colloidal properties such as mean particle diameters (Dss),distribution constants ( n ) , and sedimentation volumes (VE.) of the test suspensions prepared by this method appear in Table I. The suspensions other than the controlled ones were prepared by similar methods with appropriate modifications. After adjustment of the drug concentration in a test suspension, it was stored a t 25' until use. The syringeability for all the suspensions was good in the ) syringe ~ used in the absorption exsame needle (25G X 2.54 ~ m and periment. Procedure of Absorption Experiment-Male Wistar albino rats (250-290 g) were used in all absorption experiments. The subcutaneous region near the center of the dorsum of intact rat was selected as a model injection site and the same local clearance method described in the previous paper (5) was employed for the present investigations. Since almost all of the drug remaining in the injection site was undissolved and unchanged, the absorption experiment adopted was considered to be appropriate for understanding the true absorption phenomena from the systems of interest. Measurement of Physicochemical Properties of Test Compounds and Suspensions-For the solub es of model drugs in 0.9% (w/v) NaCl (saline) or in pH 7.25 phosphate buffer (0.067 M Na2HP04-KH2P04) d with NaC1, previously obtained data (4) were used. The es in 2% (w/v) bovine serum albumin8 in pH 7.25 isotonic phosphate buffer were determined according to a method similar to that described in the previous paper (4). The density ( p ) of the model compound, and the sedimentation volume ( Vsed) and particle size distribution (Ds8and n ) of each test suspension were also measured by the methods described in the previous report (4). Analytical Method-Samples from Absorption Experiments-The amounts of test compounds other than betamethasone dipropionate remaining in the injection site were determined colorimetrically as described previously (4). Betamethasone dipropionate was analyzed by the HPLC method under the following conditions: H P I X apparatusg; UV detector (254 nm); columnLn,25 cm X 2.1-mm 4; mobile phase, n-hexane-isopropyl ether-ethanol-HzO (5:20:1.5:10, v/v, upper layer); flow rate (pressure), 0.43-0.46 ml/min (84 kg/cm2). Samples from Other Experiments-The concentrations of model compounds other than betamethasone dipropionate in test suspensions were analyzed spectrophotometrically as described in the previous paper (4). For the analysis of betamethasone dipropionate, the optical density was read at 243 nm after dilution with ethanol-HZ0 (l:l, v/v). The sample solutions (filtrate or supernate) for the determination of the solubilities
50.0 5.0
5.0 5.0 5.0 50.0 25.0 2.5 1.25 g = -0.656 (0.049), h
Millipore memhrane filter HA., Millipore Corp., Bedford, Mass. 7 Terumo Co., Ltd., Tokyo. Sigma Chemical Co., St. Louis, Mo. Dupont 830 I L , lo Zorbax SIL,
Comment
0.27 0.14 0.025 0.015 0.12 0.10 0.046 0.032 0.020 0.027 0.045 0.11
0.10 1.00 2.00
VO:constant
CO:constant
WO:constant
= -0.315 (0.057)
0 Controlled suspension. b The values g and h were estimated by multiple regression analysis and are given together with the standard errors in parentheses.
of azo compounds in 2% bovine serum albumin solution were assayed by a colorimetric method similar to that reported previously (4). Those of sulfa drugs were assayed as follows: One milliliter of the sample solution was thoroughly mixed with 5 ml of pH 4.7 buffer [ l N HCI-1 N CH&OONa, 3 7 (v/v)] and from this mixture the sulfa drug was extracted with 10 ml of ethyl acetate. A portion of this extract was freed of the solvent in uacuo then assayed colorimetrically after diazotization according to the method described in the previous paper (4). RESULTS A N D DISCUSSION
State of the Depot a n d D r u g Absorption Kinetics-Although the subcutaneous region is composed of networks of loosely cross-linked fibrous tissues, injected solutions form a pool (depot) with lateral spread (6,7). This was also observed in experiments using aqueous suspensions of some dyes and uniform latexes. In addition, the suspension particles were confined to such networks and their spreading was not so extensive as that of the aqueous dispersion medium (vehicle) after injection. For example, even in the case of a I-ml injection of 0.5% (w/v) of uniform latex of 0.7 pm in diameter (this size seems to be the approximate lower limit for common preparations) into the dorsal subcutaneous region of rats, the resulting lateral spreading area of the particles did not exceed -30% of that of the aqueous vehicle. This spreading area became smaller with increasing particle size. These findings clearly indicated that the subcutaneously injected particles were loosely agglomerated at the injection site. In common cases, a drug in aqueous suspension injected subcutaneously must first become dissolved in the tissue fluids surrounding the particle agglomerate, diffuse into the intercellular space (or intracellular region) of the connective tissues, and then permeate the vascular membranes until it enters the blood or lymph stream. The participation of direct absorption of the fine particles by phagocytosis or some other mechanisms can not be completely denied (8,9). However, their contribution to the drug absorption may be much smaller than that of the above mentioned mechanism uia the dissolution process except for exceedingly fine particles of water-insoluble drugs (10). It was indicated previously (11) that the connective tissue ground substance formed a diffusion barrier to the subcutaneous absorption of carbohydrates with different molecular weights in aqueous solution. The relationship between the absorption rates of a variety of compounds subcutaneously implanted in rats and their physicnchemical properties Table 111-Check of Experimentally Determined g a n d h Values Using Betamethasone Dipropionate Aqueous Suspensions Experiment Number 1"
26 6
j , hr-'
CO,mg/ml
Vn, ml
2 2 1 10
0.05 0.50 0.50 0.50
j ( i ) / J ( 1) Calculated Experimental
0.48
0.49 (0.03)
0.2'2
0.26(0.03)
a Vehicle, 0.01% (w/v) polysorbate 80 + 0.01% (w/v) henzalkonium chloride + 0.38% (w/v) NaHzP042H20 1.79%(w/v)Na2HPOr12H,O; D, ( n ) ,7.1 pm ( 2 . 5 ) . Vehicle,0.5% Iw/vl methvlcellulose SM-15 0.01%( w k ) aolvsorhate 80 t 0.9% (w/v) NaCI; D.. ( n ) ,2.5 pm (2.6).
+
+
Journal of Pharmaceutical Sciences I 501 Vol. 71, No. 5, May 1982
A
B
s 0.6
-
0.4
-
+\ +\
I
I
1
0.
Figure 1-Time course of p-hydroxyazobenzene absorption from subcutaneous injection site. Each point represents the mean of a t least four experiments; vertical bar shows the standard deuiation. CO= 1 mglml; VO= 0.5 ml. were studied (2). It was concluded that the absorption from this system was solution rate limited. Visual observations of present experimental systems supported this conclusion, since the tissue fluid colored by the dissolution of a dye was almost limited to the narrow region close to the surface of the particles or their agglomerate except for a short period immediately after injection. On the basis of the above findings, it is assumed that the absorption is controlled by diffusion from the surface of the agglomerate. The process of this absorption seems to have a resemblance to that observed after intramuscular injection. Thus, the absorption model proposed for the intramuscular injection in a previous study (4) is expected to be applicable to the subcutaneous injection as well. From that model the following kinetic equations could be derived to describe the intramuscular absorption process (4). The fraction of the drug amount ( W )remaining a t the injection site to the dose ( W O was ) given as a function of time ( t ) : ( W/Wo)1l3= 1 - j t
(Eq. 1)
where j was defined as an absorption rate constant. This j could be related to the in uiuo solubility ( C s ) ,density of the drug crystal ( p ) , in uiuo dissolution rate constant ( k ) ,and the dose ( WO), using a correction parameter ( t ) for the effective surface area of the particle agglomerate for the in uiuo dissolution: j = k ~ C , p -W ~ /0~- ~ f ~ / 3
(Eq. 2)
In this equation the parameter t was assumed tobe an unknown function of the effective particle size ( D ) ,initial volume concentration (Colp) of the suspended drug, injection volume (Vo), and other factors ( U )containing hydrodynamic factors (injection speed and pressure) and the histological and physiological factors in the injection site: = F ( D , C o l p , Vo,
u)
(Eq. 3)
intramuscular absorption were checked to ascertain their applicability for the subcutaneous absorption and then developed to gain greater understanding of its phenomena. Ti m e Course of D r u g Absorption-First, the validity of Eq. 1 was checked experimentally. Figure 1 shows a plot of the residual fraction (WIWO)or its cube root of the drug remaining in the injection site against time after dorsal subcutaneous administration of the controlled aqueous suspension of p-hydroxyazobenzene into intact rats. The initial drug concentration (CO)and injection volume (Vo) are given in the legend. The cube root plot (B) gave a good linear relationship while the other (A) showed an upward curvature. This tendency was common throughout the following absorption experiments of different conditions, and the correlation coefficient obtained by the least-squares linear regression from the cube root plot was always larger than that from the linear plot. These results suggest that Eq. 1 is appropriate for describing the time course of the subcutaneous drug absorption from aqueous suspensions. In addition, the moderate magnitude of the standard deviations shown in Fig. 1demonstrated that the other factors ( U )in the parameter t might be well-controlled under the experimental conditions in this study. Effect of Particle Size on Absorption-The particle size of a suspension is an important factor for intramuscular (12-14) and subcutaneous absorption (1)as well as for absorptions from other routes (15). If the suspension particles are dispersed at the injection site too extensively to aggregate with each other, and, hence, the drug transport from the surface of each particle takes place independently, the particle size effect on the absorption may appear most strongly. tn such a case, theoretically, E = 6( W ~ / P ) ' / ~for / Dmonodispersed spherical particles (diameter, D ) and therefore is inversely proportional to the particle size (4). However, this is expected to he rare in practice because of the agglomerate formation in the injection site, except in an exceedingly dilute suspension with very fine particles. Thus, the relationship between the drug absorption
The relationship between t (or j ) and Co, VO,or D was investigated experimentally (4). In the present study, the above kinetic relationships proposed for the
0.14
-
0.10
-
0.06
-
HOURS
L.
S .1
0.02
1 I
0
'fy-.*I
I
I
4
8
12
1. 1~
PARTICLE SIZE, Dss,Mrn
F i gure 2-Effect of particle size on subcutaneous absorption of phydroxyazobenzene. Each point represents the mean of a t least three experiments; uertical bar indicates the standard deuiation. Ca = 5 rnglml; V O= 0.5 ml. 502 1 Journal of Pharmaceutical Sciences Vol. 71, No. 5, May 1982
F i g u r e 3-Relation between absorption rate constant (j) and particle size (Dss).The ualue of j was estimated by the least-squares method and is plotted with the standard error. Key: -0-, sc (VO= 0.5 ml);-0-, im (Vo = 0.05 ml);p-hydroxyazobenzene concentration of both injections, 5 mglml.
of Subcutaneous Absorption Rate Constants ( j ) with In Vitro Solubilities (6,a n d Ci) a n d Values of
Table IV-Comparison MW-0.5p-0.34
~
~
~~~~
I n Vitro Solubility C,, mg/mla C,, mg/mlb
Compound Sulfamethoxazole
5.7' 0.076 0.049 0.034 0.007 0.0003
N1-Acetylsulfamethoxazole p -Aminoazobenzene p-Hydroxyazobenzene 0 - Aminoazotoluene 1-Phenylazo-2-naphthylamine
j , hr-'
MW-O.5,,-0.34 c
7.2 <0.76 0.41 <0.69 <0.14 0.076
0.0549 0.0522 0.0672 0.0637 0.0625 0.0585
0.374 (0.015) 0.168 (0.013) 0.0652 (0.0037) 0.0420 (0.0035) 0.0193 (0.0016) 0.00241 (0.00014)
a Solubility in saline [0.9% (w/v) NaCl aqueous solution] a t 37". * Solubility.in 2% (w/v) bovine serum albumin in pH 7.25 isotonic phosphate buffer at 37". MW, molecular weight; p ! density. This term is defined in detail in the text. d Value estlmated by the least-squares method from the data shown in Fig. 6;listed with the standard error in arentheses. e Solubility in pH 7.25 isotonic phosphate buffer at 37" (solubility in saline at 37', 0.61 mg/ml). Solubilities of other test compounds in saline agreed well wit[ those in the buffer.
rate and particle size seems to be very complicated. A previous study on the intramuscular absorption showed that the increase in the absorption rate constant 0')with decreasing mean particle diameter (D,,) was remarkable in the region of D,,smaller than 2-3 pm, while it was gradual or slight in the region above this (4). Figure 2 compares subcutaneous absorption time profiles of p-hydroxyazobenzene from three preparations with different mean particle diameters (DSs)but with similar distribution constants ( n )and sedimentation volumes (Vaed):D , ( n ) ,7.4pm (2.5), 2.5pm (2.8),and 1.1 pm (2.9);V e d , 2.8-3.2cm3/g. The absorption rate constant 0') was obtained from the slope of each linear relationship in this figure and then plotted against D, in Fig. 3.For comparison, the results from similar experiments of the intramuscular route (m. gastrocnemius) are also shown. Figure 3 suggests that the relationship between j and D,, for the subcutaneous route is qualitatively similar to that for the intramuscular route. The particles smaller than 2-3 pm in diameter seem to pass more easily through the network of the fibrous tissues accompanying the spreading of the dispersion medium during injection. This may cause a looser agglomeration of the particles and result in the abrupt change in J below this particle size. Figure 4 shows the effect of particle size on the lateral spreading of the particle agglomerate. Here,six water-insoluble standard particles with different mean diameters (D, obtained from the number size distribution curve) were used for comparison. The similarity of the form of this curve to those in Fig. 3 may support the above speculation. The participation of direct absorption mechanisms involving phagocytosis must be also considered but their contributions seem to be relatively small under the present experimental conditions. In any event, the tendency of the particle size effect appears to have a very important practical meaning. Effect of Initial Dr u g Concentration and Injection Volume-If the injected particles form no agglomerate and the drug transport from each particle proceeds independently, and if this process is a rate-determining step, the absorption rate constant 0')does not depend on the dose and is inversely proportional to the particle size as mentioned previously. However, this was not the case for the subcutaneous injection of ordinary aqueous suspensions. The absorption rate constant G), given
by Eq. 2,is a function of the dose Wo and the parameter c. The term WO is the product of the initial drug concentration (CO)and injection volume ( V O ) The . parameter c is assumed to depend on COand Vo, but their relationship is not known. Therefore, the quantitative relationship between j and COor V0 is also not clear. To elucidate this relationship, the absorption time courses of the controlled suspensions of p-hydroxyazobenzene with different Co and V Owere followed, then their absorption rate constants 0') were obtained from the same plot as Fig. 1B.In Fig. 5 j is plotted against COa t fixed VO (0.5 ml) or against VOa t fixed CO(5mg/ml) on a log-log scale. The linear relationships of both plots allow the following tentative and approximate representation: where f is a constant that depends on the drug and its suspension preparation. Thus, the experimental values of g and h were estimated by multiple regression analysis from 12 sets of data for the controlled suspensions of p-hydroxyazobenzene. The original data used for this estimation and resulting g and h values are summarized in Table 11. Next, to ascertain the applicability of these experimental values (g = -0.66 and h = -0.32), additional examinations were done using aqueous suspensions of betamethasone dipropi'onate. Employing Eq. 4 and the g and h values determined above. the ratio of j ( i ) at Co(i) and Vo(i)to j(1) at Co(1) and Vo(1) for suspensions with the same drug and similar colloidal properties can be represented as:
j ( i ) / j ( I )= [Co(i)/Co(1)l-066 [ V o ( i ) l V 0 ( 1 ) ]32- ~
V,, ml 0.1
-
L '
0.1
-
0.05
-
L
.c
;.
0.5
0.2 I
I
1
1
I
1 1 1 . 1 1
I 0
1
5
10
15
MEAN PARTICLE (D), pm
20
Figure 4-Effect of particle size on the lateral spreading area o f the particle agglomerate formed by subcutaneous injection of aqueous suspension. Each point represents the mean of three experiments; vertical bar indicates the standard deuiation. These data were obtained using water-insoluble standard particles of uarious sizes under the following conditions: CO= 5 mglml; VO = 1.0 ml; measurement, 5 min after injection.
0.01
L r
2 I
5 I
1
1
1
1
I
,
-\
0.021 0.6
(Eq. 5)
Table I11 compares the calculated and experimental ratios j(i)/j(l)for the case where Co(i)/Co(l)= 10 or Vo(i)/Vo(l)= 10.These comparisons showed good mutual agreements, despite the fact that the vehicle and the colloidal properties of the suspensions tested here differed considerably from those of the controlled suspension of p-hydroxyazobenzene used for the estimation ofg and h values. Therefore, Eq. 5,obtained ex0.05
Q
(Eq. 4)
j = fC&Voh
' ' 0.5
I
t " 1 '
1
2
5
10
20
I
1
,
50
C,. mglrnl
Figure 5-Relation between absorption rate constant 6) and injection volume (Vo) or initial drug concentration (Co) for t h e controlled suspension of p-hydroxyazobenzene. T h e value of j U I Q S estimated by the least-squares method. Key: - 0 - , constant VO(0.5 ml);-0-, constant CO(5mglml). Journal of Pharmaceutical Sciences I 503 Vol. 71, No. 5. May 1982
HOURS
- 10
1.
lo-'
.
-1
"
"
L
L
r
c
..
'\
- lo-'
.
-
- /oO0
10-2
-
lo-'
lo-'
IO-~
lo-*
10
1
Ci,mg/ml
Figure 6-Comparison of absorption rate among uarious compounds (controlled suspension). Each point represents the mean of three or four experiments; vertical bar indicates the standard deuiation. Co = 5 mglml; VO = 0.5 ml. Key: -@-, I-phenylazo-2-naphthylamine; -A-, o-aminoazotoluene; p-hydroxyazobenzene; -0-, paminoazobenzene; -A-, N'-acetylsulfamethoxazole; -0-, sulfamethoxazole.
Fi g u r e 7-Relationship between absorption rate constant (j) a nd in vitrosolubility (Cs').The ualue of j was estimated by the least-squares method from the data shown in Fig. 6. Key: -@-, sc (C, = 5 mglml; vo = 0.5 ml); -0-, im (Co = 5 mglml; Vo = 0.05 ml), cited from a previous study ( 4 ) .
perimentally, is expected to be generally applicable for aqueous suspensions other than the model ones tested here. Using the above results, the relationship between the parameter t and COor VOcan be clarified empirically in the following manner:
As discussed previously, the absorption is inferred to be a diffusioncontrolled process. Diffusion coefficients of compounds having low molecular weight (MW) are known to be inversely proportional to the square root of MW (16). The in uiuo dissolution rate constant ( k ) in Eq. 9 is considered to be proportional to the diffusion coefficient, like that in uitro (2,17). Therefore, Eq. 9 can be converted to
-+-,
j = fCO-O66VO-0.32
(Eq. 6)
Employing the relationship WO= CoVo, Eq. 2 can be rewritten as:
j =
(Bd/3)MW-0.5p-0.34C,Co-0.66V
-0.32
(Eq. 11)
where B is a constant. Thus, the values of MW-o.5p-o.34for model drugs were calculated and are in Table IV as the correction term. However, From the equivalency of Eqs. 6 and 7, the parameter t must satisfy the these values were similar to each other for the compounds tested here. following relationship: In contrast, the in uitro solubility was a more dominant factor and had a positive relation with the absorption rate constant. c = F(D, Colp, VO,U ) = ~ ( C O / ~ ) - ~ . ~(Eq. ~ V8 )~ ~ . ~Equation ~ 11 means that the plot of j (or J,,,, = jMW0.5e0.34,if the correction for MW and p is necessary) against in uiuo solubility (C,)'on where 6 is a term that depends on the remaining factors D and U . a log-log scale gives a straight line with a slope of unity a t the constant Equation 8 means that the parameter t depends little on the injection initial drug concentration (CO)and injection volume (VO).The closed volume ( V O )but decreases by increasing initial drug concentration (CO). circles in Fig. 7 show this plot using in uitro solubility (C',) in Table IV This also implies that the degree of agglomeration may tend to increase instead of in uiuo solubility (Cs). This gave a nearly straight line with a with an increase in Co. Substitution of Eq. 8 for c in Eq. 7 yields: slope of 0.53 (regression equation, log j = 0.525 log Cb - 0.578; correlation coefficient, r = 0.956). Using j,,,, in place of j , a similar relationship was j = (kC,j/3)p-0.34CO-0.66VO-0.32 (Eq. 9) also obtained (log jcorr = 0.534 log C', 0.660, r = 0.953). The above empirical equation obtained for the subcutaneous route In Fig. 7, similar data for the intramuscular administration (m. gasdemonstrates a great similarity to that for the intramuscular route, which trocnemius), cited from a previous study (4), are also plotted with open was given by the following equation (4): circles. Remarkably, the slopes of these plots for the two routes were almost identical. This shows a strong similarity in the absorption mechaj , = (k,C,6,/3)p-0.45Co-0.55V~-o 32 (Eq. 10) nism between the two routes. The deviation of the slope from unity obIt should be noted that the correlation between the absorption rate served in Fig. 7 may he attributed mainly to a gradual increase in the ratio constant and injection volume is identical for the two routes. C,/C, with decreasing C; caused by the relatively larger. solubilization Although Eqs. 9 and 10 are derived empirically, they should be very effect of the protein components in the body fluid for the drug having important and useful since they give relationships between the dose and a smaller solubility. The comparison of C', and C ; in Table IV supports absorption rate constant which have been unknown until now. this explanation. Comparison of Absorption R a t e among Various CompoundsThe good linear relationship between log j (or log j,,,,) and log C; Equation 9 shows that the absorption rate constants (j) of different mentioned previously is expected to be applicable for predicting the compounds depend upon the values of k , C,, and p when the preparasubcutaneous absorption rates of other compounds in the controlled tion-dependent parameters (6) are the same. As defined previously, p , suspensions from C,. For instance, the prediction (from the regression k , and C, represent the density of the drug, the in uiuo dissolution rate equation, log j,,, = 0.534 log C; 0.660) for the test material, betamconstant, and in uiuo solubility in the injection site, respectively. Of these ethasone dipropionate (MW = 504; p = 1.23 g/cm3; C, = 0.002 mg/ml) factors, only the value of p can be determined by an in uitro experiment; gave a value of 0.007 hr-I for j (at CO= 5 mg/ml and VO= 0.5 ml), which the other two ( k and C,) cannot be completely estimated from such an almost agreed with the experimental value of 0.0096 (f0.0011) hr-' obexperiment. From a practical point of view, an attempt was made to served using the following controlled suspension: D,,= 4.3 pm; n = 2.6. clarify the quantitative relationship between j and the above three factors This satisfactory result suggests that similar prediction will be possible for other drugs. (or their first approximations). Figure 6 compares the subcutaneous absorption time profiles of six test Comparison of Absorption Rates between Subcutaneous a n d compounds. In this comparison, the controlled suspensions with similar I n t r am u scu l ar Routes-Hitherto, drug absorption kinetics and colloidal properties (Table I) were used to minimize the variation in 6 in mechanisms from aqueous suspensions in the subcutaneous route have Eq. 9. All the compounds gave good linear relationships with different been clarified, and their strong similarity to those by the intramuscular slopes. Their observed absorption rate constants (j) are summarized in route, discussed previously (4), have been pointed out. A direct comTable IV with their physicochemical factors such as in uitro solubilities parison of absorption rates between the two routes seems to be usable but there has been little available data (18). Table V shows this comin saline and 2% (w/v) bovine serum albumin (C, and C;, respectively). j = kC,1'-2/3~C~-1/3V~-1/3/3
(Eq. 7)
+
+
504 I Journal of Pharmaceutical Sciences Vol. 71, No. 5,May 1982
Table V-Comparison of Absorption Rate Constants ( j ) between Subcutaneous (sc) and Intramuscular (im) Routes a j , hr-I
Compound Sulfamethoxazole p-Aminoazobenzene p-Hydroxyazobenzene o-Aminoazotoluene 1-Phenylazo-2-naphthyl-
sc
imc
0.78 0.14 0.090 (0.12 f 0.02)d 0.040 0.0050
1.10 f 0.07 0.18 f 0.01 0.17 f 0.01 0.093 f 0.005 0.0093 f 0.0006
smine
*
Controlled suspension.CO,5 mg/ml; VO,0.05 ml. Estimated by extrapolation of data shown in Table IV usin Eq 5 Experimental data (with standard error) cited from the previous reportT4). d Experimentalvalue (with standard error). parison using five controlled suspensions. To compare a t the same drug , values estimated by concentration (CO)and injection volume ( V O )the extrapolation of the data shown in Table IV using Eq. 5 were used for the absorption rate constants 0’) in the subcutaneous route. This comparison shows that the absorption rate from the subcutaneous route is slower than that from the intramuscular route for all the test suspensions. A similar tendency was previously observed for injections of drug-oil solutions (5). The relationship between j and COin the subcutaneous route differed slightly from that in the intramuscular route (Eqs. 9 and 10). Therefore, it should be noted that the difference in j shown in Table V may increase with increasing CO. REFERENCES (1) V. G. Foglia, J. C. Penhos, and E. Montuori, Endocrinology, 57, 559 (1955). (2) B. E. Ballard and E. Nelson, J. Pharmacol. Erp. Ther.,135,120 (1962). (3) B. E. Ballard and E. Nelson, J. Pharm. Sci., 51,915 (1962).
(4) K. Hirano, T. Ichihashi, and H. Yamada, Chem. Pharm. Bull., 29,817 (1981). (5) K. Hirano, T. Ichihashi, and H. Yamada, J . Pharm. Sci., 71,495 (1982). (6) E. A. Brown, T. G. Metcalf, and L. W. Slanetz, Ann. Allergy, 19, 1016 (1961). (7) F. L. Ashley, S. Braley, T. D. Rees, D. Goulian, and D. L. Ballantyne, Jr., Plastic Reconstruc. Surg., 39.411 (1967). (8) C. R. Beresford, L. Golberg, and J. P. Smith, Br. J.Pharmacol., 12, 107 (1957). (9) T. D. Rees, D. L. Ballantyne, Jr., I. Seidman, and G. A. Hawthorne, Plastic Reconstruc. Surg., 39,402 (1967). (10) J. Lewin and F. Huidobro, Acta Physiol. Lat. Am., 3, 17 (1953). (11) E. Secher-Hansen, H. L. Langgard, and J. Schou, Acta Pharmacol. Toxicol., 26,9 (1968). (12) F. H. Buckwalter and H. L. Dickison, J . Am. Pharm. Assoc. Sci. Ed., 47,661 (1958). (13) L. G. Miller and J. H. Fincher, J . Pharm. Sci., 60,1733 (1971). (14) N. Kitamori, S. Kawaziri, and T. Matsuzawa, “Abstracts of Papers,” 93th Annual Meeting of Japan Pharmaceutical Society, Tokyo, April 1973, p. 266. (15) J. H. Fincher, J . Pharm. Sci., 57,1825 (1968). (16) W. D. Stein, “The Movement of Molecules across Cell Membranes,” Academic, New York, N.Y. 1967, p. 67. (17) W. Nernst, 2. Phys. Chem., 47,52 (1904). (18) J. J. Nora, W. D. Smith, and J. R. Cameron, J . Pediatr., 64,547 (1964). ACKNOWLEDGMENTS The authors thank Mrs. J. Kagawa for her technical assistance and Prof. M. Nakagaki, Kyoto University, for his valuable comments on the manuscript.
Dissolution and Bioavailability Studies of Whole and Halved Spstained-Release Theophylline Tablets KEITH J. SIMONSX*,EVELYN M. FRITH*, and F. ESTELLE R. SIMONS$ Received July 24,1980, from the ‘Faculty of Pharmacy, and f Section of Allergy and Clinical Immunology, Department of Pediatrics, Facultv Accepted for publication August 5, 1981. of Medicine, University of Manitoba, Winnipeg, Manitoba, Canada, R3T 2N2. Abstract 0 In dissolution studies of whole and halved 100-mg sustained-release theophylline tablets, drug release from halved tablets was significantly higher. These differences were not reflected in the bioavailability studies. The area under the curve (AUC) mean absorption time and fraction-of-dose recovered in urine at 24 hr were not significantly different following the ingestion of whole or halved 100-mg tablets. The elimination rate constant, half-life, volume of distribution, plasma, and renal clearance values were consistent with values reported previously. Discrepancies were found in the 24-hr metabolite distribution as compared to literature values and may be accounted for by the age and health of the subjects and the frequency of dosing.
phylline tablets’ were used to study the effect of halving tablets on dissolution and bioavailability. No published information about the dissolution of these tablets was available. After oral administration of the 100-mg tablet, however, 90% of the dose was absorbed within 14 hr and almost 100% was absorbed by 28 hr (1). When 300-mg tablets were dissolved, 50% of the dose entered solution by 2 hr and > 90% of the dose entered solution by 6 hr (1).
Keyphrases 0 Dissolution-whole and halved sustained-release theophylline tablets Sustained-release system-dissolution of whole and halved theophylline tablets 0 Bioavailability-whole and halved sustained-release theophylline tablets 0 Theophylline-bioavailability and dissolution study of whole and halved sustained-release tablets
EXPERIMENTAL
Breaking sustained-release theophylline tablets in half is commonly practiced to achieve more accurate milligrams per kilogram dosing in children. The extent to which this affects dissolution and bioavailability is unknown. In this investigation, 100-mg sustained-release theo0022-354918210500-0505$01.0010 @ 1982. American Pharmaceutical Association
Dissolution-The official USP dissolution apparatus was used (2). Simulated gastric and intestinal fluids were used as dissolution media (2). Simulated gastric fluid, USP (Z),was prepared by dissolving 2 g of sodium chloride and 3.2 gof pepsin in 7 ml of HCI and diluting the solution to 1000 ml with distilled water. This test solution had a pH of 1.2, Simulated intestinal fluid, USP (2), was prepared hy diswlv~ng6.8 g Theo-Dur, Astra 1M4.
Pharmaceuticals Canada Ltd.. Mississauya, Canada IAX
Journal of Pharmaceutical Sciences I 505 Vol. 71,No. 5,May 1982